LOCUS       sequence-328774-        6511 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression plasmid for His6-MEK1C218.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6511)
  AUTHORS   Lai KY, Galan SRG, Zeng Y, Zhou TH, He C, Raj R, Riedl J, Liu S, 
            Chooi KP, Garg N, Zeng M, Jones LH, Hutchings GJ, Mohammed S, Nair 
            SK, Chen J, Davis BG, van der Donk WA
  TITLE     LanCLs add glutathione to dehydroamino acids generated at 
            phosphorylated sites in the proteome.
  JOURNAL   Cell. 2021 Apr 27. pii: S0092-8674(21)00436-0. doi: 
            10.1016/j.cell.2021.04.001.
  PUBMED    33932340
REFERENCE   2  (bases 1 to 6511)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell. 2021 
            Apr 27. pii: S0092-8674(21)00436-0. doi: 
            10.1016/j.cell.2021.04.001."
COMMENT     Sequence Label: MEK1C218
FEATURES             Location/Qualifiers
     source          1..6511
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     67..86
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        67..85
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    86..110
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             125..147
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             166..183
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             193..210
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1442..1459
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1512..1530)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1526..1573
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1610..2065
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1697..1716)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1907..1928
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2157..2972)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2157..2972)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2880..2899
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2964..2983
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3094..3682
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3583..3602
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3836..3853
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3868..4007
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3933..3952)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4093..4115
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4109..4300)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5109..6191)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     6153..6172
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6192..6269)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6475..6494
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg
       61 cgaaattaat acgactcact ataggggaat tgtgagcgga taacaattcc cctctagaaa
      121 taattttgtt taactttaag aaggagatat accatgttca gcagccatca tcatcatcat
      181 cacagcagcg gcctggtgcc gcgcggcagc catatgccga aaaaaaaacc gaccccgatt
      241 caactgaacc cggctccgga tggctccgct gtcaatggca cctcaagcgc agaaacgaac
      301 ctggaagcgc tgcagaaaaa actggaagaa ctggaactgg atgaacagca acgtaaacgc
      361 ctggaagcct ttctgaccca gaaacaaaaa gtgggcgaac tgaaagatga cgatttcgaa
      421 aaaatcagtg aactgggcgc cggtaacggc ggtgtggttt ttaaagtcag ccataaaccg
      481 tctggtctgg tgatggcacg taaactgatt cacctggaaa tcaaaccggc tattcgtaac
      541 cagattatcc gcgaactgca agtgctgcat gaatgcaatt ctccgtatat tgttggcttt
      601 tatggtgcgt tctacagtga cggcgaaatt tccatctgta tggaacacat ggacggcggt
      661 agcctggatc aggttctgaa aaaagcaggc cgtatcccgg aacaaattct gggtaaagtc
      721 tctattgctg tgatcaaagg cctgacgtac ctgcgtgaaa aacataaaat catgcaccgc
      781 gatgtgaaac cgtcaaacat cctggttaat tcgcgcggtg aaattaaact gtgcgacttt
      841 ggcgttagcg gtcagctgat tgattgtatg gcgaactctt tcgtcggcac ccgtagttat
      901 atgtccccgg aacgcctgca gggtacgcat tactcagtgc aatcggatat ctggtcaatg
      961 ggcctgtcgc tggttgaaat ggccgttggt cgttatccga ttccgccgcc ggacgccaaa
     1021 gaactggaac tgatgtttgg ctctcaggtt gaaggtgatg cggccgaaac cccgccgcgt
     1081 ccgcgtaccc cgggtcgtcc gctgagctct tatggtatgg acagccgtcc gccgatggca
     1141 atcttcgaac tgctggatta cattgtgaat gaaccgccgc caaaactgcc gagcggcgtg
     1201 tttagcctgg aatttcagga cttcgtcaac aaatgtctga tcaaaaatcc ggcagaacgt
     1261 gctgatctga aacaactgat ggtccacgct tttattaaac gctccgacgc ggaagaagtg
     1321 gatttcgccg gttggctgtc ttcgaccatt ggcctgaatc aaccgtcaac gccgacccac
     1381 gctgctggtg tgtaaggatc cgaattcgag ctccgtcgac aagcttgcgg ccgcactcga
     1441 gcaccaccac caccaccact gagatccggc tgctaacaaa gcccgaaagg aagctgagtt
     1501 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
     1561 gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt
     1621 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     1681 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     1741 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     1801 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     1861 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     1921 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     1981 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     2041 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
     2101 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaat taattcttag
     2161 aaaaactcat cgagcatcaa atgaaactgc aatttattca tatcaggatt atcaatacca
     2221 tatttttgaa aaagccgttt ctgtaatgaa ggagaaaact caccgaggca gttccatagg
     2281 atggcaagat cctggtatcg gtctgcgatt ccgactcgtc caacatcaat acaacctatt
     2341 aatttcccct cgtcaaaaat aaggttatca agtgagaaat caccatgagt gacgactgaa
     2401 tccggtgaga atggcaaaag tttatgcatt tctttccaga cttgttcaac aggccagcca
     2461 ttacgctcgt catcaaaatc actcgcatca accaaaccgt tattcattcg tgattgcgcc
     2521 tgagcgagac gaaatacgcg atcgctgtta aaaggacaat tacaaacagg aatcgaatgc
     2581 aaccggcgca ggaacactgc cagcgcatca acaatatttt cacctgaatc aggatattct
     2641 tctaatacct ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca tgcatcatca
     2701 ggagtacgga taaaatgctt gatggtcgga agaggcataa attccgtcag ccagtttagt
     2761 ctgaccatct catctgtaac atcattggca acgctacctt tgccatgttt cagaaacaac
     2821 tctggcgcat cgggcttccc atacaatcga tagattgtcg cacctgattg cccgacatta
     2881 tcgcgagccc atttataccc atataaatca gcatccatgt tggaatttaa tcgcggccta
     2941 gagcaagacg tttcccgttg aatatggctc ataacacccc ttgtattact gtttatgtaa
     3001 gcagacagtt ttattgttca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     3061 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     3121 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     3181 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     3241 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     3301 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     3361 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     3421 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     3481 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     3541 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     3601 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     3661 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     3721 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     3781 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     3841 gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg
     3901 cggtatttca caccgcaatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3961 agccagtata cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc
     4021 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     4081 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     4141 cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct
     4201 gttcatccgc gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa
     4261 agcgggccat gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg
     4321 gatttctgtt catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg
     4381 ttactgatga tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat
     4441 ggatgcggcg ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag
     4501 atgtaggtgt tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg
     4561 tgcagggcgc tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc
     4621 atgttgttgc tcaggtcgca gacgttttgc agcagcagtc gcttcacgtt cgctcgcgta
     4681 tcggtgattc attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg
     4741 acaggagcac gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg gcctgcttct
     4801 cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc
     4861 cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga
     4921 aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca
     4981 taagtgcggc gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg
     5041 ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg
     5101 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     5161 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc
     5221 accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc
     5281 aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc
     5341 gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca
     5401 acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca
     5461 accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg
     5521 gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga
     5581 tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac
     5641 agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct
     5701 tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc
     5761 ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta
     5821 atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg
     5881 acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat
     5941 ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca
     6001 atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc
     6061 tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc
     6121 accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt
     6181 actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg
     6241 cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc
     6301 ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa
     6361 tggtgcatgc aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc
     6421 cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc cgatcttccc catcggtgat
     6481 gtcggcgata taggcgccag caaccgcacc t
//
